A blood coagulation mechanism is broadly divided into two pathways in general. One is an intrinsic pathway, which is initiated by contact activation of blood coagulation factor XII with a foreign substance, and finally produces thrombin through multistep reactions, and the other is an extrinsic pathway, which is initiated by activation of blood coagulation factor X with blood coagulation factor VII and tissue thromboplastin, and produces thrombin in the same manner as described above (FIG. 1). In both of the pathways, coagulation eventually occurs by conversion of fibrinogen to fibrin through an action of produced thrombin. In order to clarify the presence or absence of abnormality or a cause of the abnormality in such blood coagulation mechanism, some blood coagulability tests using a blood coagulation activator are available and widely used in actual clinical examinations.
The blood coagulation activator and the blood coagulability tests using the same are as follows:
1) Blood Coagulability Tests Using Thrombin
a fibrinogen measurement, an ATIII measurement, and a thrombin time measurement;
2) Blood Coagulability Tests Using Tissue Thromboplastin
a prothrombin time measurement, an activity measurement of factor II, V, VII, or X using the prothrombin time measurement, and a measurement of complex factors (a thrombo test, a hepaplastin test, and the like); and
3) Blood Coagulability Tests Using Phospholipids
a partial thromboplastin time measurement, an activated partial thromboplastin time measurement, an activity measurement of factor VIII, IX, XI, or XII, prekallikrein, or high molecular kininogen using the activated partial thromboplastin time measurement, a viper venom time measurement, a quantitative determination of factor X using the viper venom time measurement, a lupus anti-coagulant (LA) measurement using a diluted viper venom time measurement, a protein C activity measurement, and a thromboplastin generation test.
In any of the above-mentioned tests 1) to 3), a time from initiation of coagulation by mixing a reagent including a blood coagulation activator and the like with a specimen from a patient to final conversion of fibrinogen into fibrin to be deposited is measured.
Methods of detecting coagulation in the blood coagulability test can be broadly divided into a mechanical detection method and an optical detection method. The mechanical detection method refers to a method including: monitoring by a magnetic force or the like a magnetic substance or the like loaded into a reaction solution; and detecting a decrease in mobility of the magnetic substance due to an increase in viscosity by coagulation. The optical detection method refers to a method including detecting white turbidity of a reaction solution due to coagulation as a change in transmitted light or scattered light, and is most widely used because the method is relatively simple. These are generally detected by an automated analyzer at present. One example of a change curve for scattered light intensity obtained in a scattered light detection method is shown in FIG. 2. In the figure, the point A indicates a point at which the coagulation is initiated by mixing a reagent including a blood coagulation activator, subsequently fibrin deposition is initiated through multistep reactions, and as a result, a change in scattered light intensity appears after the point B. Fibrinogen is in turn consumed and almost depleted in the reaction solution. Then, the scattered light intensity exhibits no change, the curve becomes flat as shown at the point C, and the coagulation is terminated. A coagulation time is calculated with known calculation parameters based on such change curve for the scattered light intensity (Patent document 1). Here, it is obvious that when a maximum of the change in scattered light intensity is defined as ΔH, as the ΔH is larger, the coagulation is detected more sharply and more correct detection becomes possible. Thus, the reagent for measuring blood coagulability desirably has a property in which an optical change is largely displayed. However, when the change from the point A to the point C occurs in an extremely short period of time, it is impossible to measure a coagulation time with good accuracy. Thus, a substance for prolonging the coagulation time is generally added to adjust the coagulation time to a desired time. A blood coagulation time prolonging agent is exemplified by halide salts of alkali metals and alkali earth metals, including sodium chloride in Patent document 2, and by sodium propionate in Patent document 3. However, there has been a problem in that such conventional blood coagulation time prolonging agent also has a large negative effect of reducing ΔH in an addition dose-dependent manner.
For the purpose of solving the problems in those conventional methods, there is proposed a method including using a reagent supplemented with a high molecular material such as polyethylene glycol, polyvinyl alcohol or high molecular polysaccharide (Patent documents 4 and 5).